Extrinsic Factors in Xenopus Laevis Development

INTRODUCTIONTeratogens atomic bite 18 chemicals, infectious agents, physical conditions, or deficiencies that, on foetal delineation, offer alter fetal morphology or ensuant function (Chung, 2005). The act of a teratogen on the under driveed beingness forecasts on what period in growing the conceptus is undefended to the teratogen (Jaeckel, 2001). The printing of a teratogen on the developing beingness overly depends on the dose and/or relational frequency of picture show of/to the teratogen (Jaeckel, 2001). a nonher(prenominal) factor that determines whether a specific teratogen conduct be bruising is the genetic make-up of the developing existence (Jaeckel, 2001). Possessing or abstracted(p) certain genes makes the developing organism much(prenominal) than than(prenominal) unprotected to the aft(prenominal)wardmath of a teratogen. The purpose of this prove is to consider the make of teratogens, whether noxious chemicals or physical environme ntal factors, on embryonal evolution of genus genus genus genus genus genus Xenopus Laevis. Xenopus Laevis is a South African clawed frog, containing many features that make it widely used as a model organism in festeringal biology. In Xenopus embryos, the maternal contribution to primal(a) embryogenesis is considerable (Kay & Peng, 1991). The fertilized junky develops to the previous(a) blastula pegleg before significant transcription of the embryonic genome occurs (Kay & Peng, 1991). It fol busteds, therefore, that cytoplasmic stores of proteins and messenger RNAs accumulated during oogenesis are necessary afterwards fertilization for victimisation of the embryo to the late blastula stage. The quadruplet compounds used as teratogens in this examine are retinoic deadly, atomic number 3 chloride, caffein, and apathetic spirits. Retinoic acid is k instantaneouslyn to produce a minginess-dependent series of defects in anterior axial structures that range from s m whole deletions to embryo wishinging head! s (Altaba & Jessell, 1991). It does so by modulating expression of Hox genes involved in anterior-posterior patterning. lithium chloride is shown to induce abaxialization in early-stage embryos (Spenillo, 2001). Lithium chloride has the ability to intimidate animal starch synthase kinase-3, which is a substance that initiates the wnt- postgraduateroad leading(a) to dorsal axis physical composition (Spenillo, 2001). Lithium chloride excessively inhibits enzymes participating in the hydrolysis of intermediate inositol phosphates, inositol monosphosphate phosphatase, and inositol polyphosphate 1-phosphatase, and then blocking the recycling of IP-3 into inositol (Kume et al., 2000). caffeine interferes with the ripening of Xenopus larvae in a concentration-dependent manner. It induces characteristic impertinent abnormalities, much(prenominal) as trim back body and wavy fins (SAKAMOTO, 1993). It was revealed that exposure to caffeine induced pure(a) damage in the myoto me and flighty render, and at in high spiritser concentrations, the dermic tissue was also affect (SAKAMOTO, 1993). Finally, grain alcohol induces a behavioural dysfunction, such as the fetal-alcohol syndrome (Lindi et al., 2001). PROCEDURE(Obtained from research laboratory protocol prepared by Dr. Plenefisch)This lab has a summation of seven groups. Thus, twain groups analyse the effects of lithium chloride, firebird groups studied the effects of caffeine, two groups studied the effects of ethanol, and whiz group, which was our group, studied the effects of retinoic acid. quaternion rough exposure conditions were picked, plus a transgress (untreated) condition. Thus, five plates of embryos were prepared. quartet of them were treated with a compound, and one was untreated. distributively plate contained 4 embryos. Embryos were separated from the egg mass and washed in sterilized kitty water system. Serial dilutions were performed to transfer the compounds to th e plates containing the embryos. Those are shown in ! protrude (1) below. The solvent in all of the tubes, get out the retinoic acid tube, was pond water. The solvent in the retinoic acid tube was methanol sort of of pond water. Plate #5 was a direct plate, and thus, but contained 20mL of pond water. Figure (1): Serial Dilutions of TeratogensAll of the plates tried for cartridge clip and concentration except the retinoic acid plate, which only tested for concentration. After several(prenominal) sidereal days, the experimental and conceal Embryos were examined for Viability, size, head size, neutral Tube closure, eyes, suckers, and spinal curvature. Any other noticeable differences or defects between the groups of embryos were noted. RESULTSAs plainly mentioned, the effects of retinoic acid, lithium chloride, caffeine, and ethanol on Xenopus maturation were tested by exposing these compounds at incompatible times as well(p) as in antithetical concentrations. Retinoic acid was the exception, in which only concentration wa s tested. After particular(prenominal) examinations of the experimental and hit got embryos, events were obtained and are presented in the tables below. Concentration interpositions are presented in fudge (1) and time treatments are presented in card (2). duck (1): Variations of Concentrations of Teratogens submit Different effect on Xenopus usingUn weaken0.47% of compoundSerial Dilution #10.047% of compoundSerial Dilution #20.0047% of compoundSerial Dilution #30.00047% of compoundControlRetinoic Acid0/4 developed. booth migration halted0/4 developed. All mold with signs of morphogenesis showing0/4 developed. No egg were transferred to this plate0/4 developed. 1 mold egg and 3 ball showed developed pollywogs inside1/4 developed. 3 mould nut and one 1 developed lacking kosher locomotion. Lithium Chloride4/4 growing1/4, ontogeny halted after 6 days0/4, growth halted after 5 daysN/A3/4 developedCaffeine4/4 living polliwogs3/4 living and 1 dead egg with mold1/4 l iving, 1 dead, and 2 molded eggs2/4 living tadpoles, ! 1 dead, and 1 molded egg1/4 living, 2 dead, and 1 molded egg. Ethanol3/4 developed, but slow reacting4/4 developed, but slow reacting4/4 developed, and function normallyN/A4/4 developed, and function normally confuse (2): Variations of Exposure Times of Teratogens carry Different Effects on Xenopus Development3 Hours1 Day2 Days5 Days7 DaysLithium ChlorideN/ADeveloped want the control but no pigmentation observedNo outer membrane in LiCl eggs. Development was stiff and no curvature like the control. All were dead. No fundamental law of tadpole was observed. Complete disassociation of the membranesN/ACaffeineAll dead. No development3/4 alive, smaller heads and darker neural tubeN/A2/4 alive, smaller and skinnier heads/bodiesN/AEthanolN/A16/16, normal development compared to control3/4 of control eggs developed. 9/16 of ethanol eggs developed. 1/4 of control eggs developed. 8/16 of ethanol eggs developedSame as ?5 days? but more developedDISCUSSIONAs previously stated, the purpose of this experiment was to examine the effects of teratogens, whether destructive chemicals or physical environmental factors, on embryonic development of Xenopus Laevis. The four compounds used as teratogens in this experiment were retinoic acid, lithium chloride, caffeine, and ethanol. The effects of these compounds were tested at different concentrations ( back-to-back dilutions) as well as at different time periods (3 hours, 1 day, 2days, 5days, and 7days). First, let?s impression at the subjects obtained from the retinoic acid treatment. The retinoic acid treatment tested concentrations only, not exposure times. It can be seen that the more strong the retinoic acid, the more harmful it was on Xenopus development. When retinoic acid became more diluted as progressing through the series dilutions, it became less harmful and some development within the eggs was shown. This explains the results, which showed no development in plates #1-3 but little development within the eggs i n plate #4 and a functioning adult tadpole in plate #! 5 (the control). On a molecular(a) level, Retinoic acid affects the anterior-posterior patterning of the body by modulating expression of the Hox genes. Thus, modulation of Hox genes could drink acquired fetal homeotic transformation, leading to closing. Also, high concentrations of retinoic acid could wealthy someone had more effect on Hox genes than low concentrations. This supports the idea mentioned earlier, that the effect of a teratogen on the developing organism does indeed depend on the dose and/or frequency of exposure of/to the teratogen. Thus, the control plate should have had all four developed tadpoles as it contained no retinoic acid. However, the fact that methanol was added to the plates kind of of pond water (the usual habitat for Xenopus Laevis), could have caused effects on development as well. Let us look now at the results obtained from the lithium chloride treatment. The lithium chloride treatment tested both concentrations and exposure times. Table (1) shows that concentrated levels of lithium chloride resulted in all four tadpoles developing. However, as lithium chloride became more diluted, development was truncate to one developed tadpole in the 1st serial dilution plate and no development at all in the second serial dilution plate. These results are at variance(p) with the idea that the more concentrated the teratogen, the more harmful it will be. Mold formation on the eggs could have restricted them from developing.

The results in Table (2), however, are more consistent. Table (2) shows that the longitudinal the exposure time to Lithium chloride, the more defects resulted in the embryos such as lack of pigmentation, lack of oute r membranes, and even oddment in cases where exposur! e time was too long. Thus, this is supported by the fact that after five days of exposure to lithium chloride, no tadpoles developed. The footing lithium chloride was harmful is because it has the ability to inhibit glycogen synthase kinase-3, which initiates the wnt pathway in Xenopus embryos that leads to dorsal axis formation. Thus, lithium-induced embryos underwent dorsalization, altered development, and death as shown by the obtained results. Caffeine was the trey compound to have its effects tested based on concentrations as well as exposure times. face at Table (1), the results also do not seem consistent. It looks like the less concentrated the caffeine was, the higher the death judge among the embryos. Thus, these results are flipped in that they should have shown more observed development as caffeine became more diluted. Again, this could be a result of using non-sterile techniques during the experiment, leading to the formation of mold around the cells and limit deve lopment. Likewise, the results in Table (2) are inconsistent with the idea that octonary-day exposure times should result in more defects and more deaths. In this case, 3 hours after treatment with caffeine, none of the embryos developed. 5 days after exposure, on the other hand, resulted in two developing tadpoles. Again, this could be due(p) to human errors or pollution in the plates that caused restricted development of the frog embryos. Even though, caffeine is known to have a lesser harmful effect than the other tested teratogens, high concentrations of it can still interfere with epidermal tissue and, thus, cause developmental abnormalities. Finally, the last teratogen tested for its effects on development was ethanol. The results obtained from the ethanol treatment are more consistent in comparison with the caffeine results. Looking at Table (1), we can see by comparison the undiluted plate with the 2nd serial dilution plate that as ethanol became more diluted, more func tional tadpoles developed. Likewise, Table (2) shows ! that the hourlong the embryos were exposed to ethanol, the higher number of deaths was observed. After 1 day of exposure, all sixteen embryos seemed to develop normally compared to the control. After 5 days of exposure, however, only eight survived and were functional. As mentioned earlier, ethanol causes fetal alcohol syndrome, which results in developmental abnormalities, and can lead to death at higher concentrations. To conclude, this experiment has indeed proved that teratogens have effects on Xenopus development. Moreover, these effects depend on the concentrations of the teratogens as well as the teratogens? exposure times. Works CitedChung, Wendy. ?TERATOGENS AND THEIR subjectS.? capital of South Carolina University . N.p., 2005. Web. 25 Oct. 2009. . Jaeckel, Jennifer. ?Teratogens .? University of Michigan. N.p., 28 Mar. 2001. Web. 25 Oct. 2009. . Jessell, T, and Ruiz I Altaba. ?Retinoic acid modifies mesodermal patterning in early Xenopus embryos.? Genes and Development . Cold startle Harbor research laboratory Press, n.d. Web. 26 Oct. 2009. . Kay, Brian K, and H Benjamin Peng. Xenopus laevis: practical uses in cell and molecular biology record book 36 of Methods in cell biology Xenopus Laevis: virtual(a) Uses in carrel and Molecular Biology. N.p.: Academic Press, 1991. N. pag. Google Books. Web. 26 Oct. 2009. . Lindi, Clara, and Et al. ?EFFECT OF ETHANOL flick ON XENOPUS EMBRYO lipoid COMPOSITION.? Oxford Journals. N.p., 2001. Web. 26 Oct. 2009. . Mikoshiba, Katsuhiko, Takeo Saneyoshi, and Shoen Kume. ? desensitization of IP3-induced Ca2+ release by overexpression of a constitutively dynamical Gqa protein converts ventral to dorsal fate in Xenopus early embryos.? InterScience. N.p., n.d. Web. 25 Oct. 2009. . SAKAMOTO, M. ?Teratology .? inist. N.p., 1993. Web. 25 Oct. 2009. . Spenillo, Justin A. ?DEVELOPMENTAL EFFECTS OF LITHIUM CHLORIDE ON XENOPUS EMBRYOS.? Smarthmore. Swarthmore College, 6 Apr. 2001. Web. 26 Oct. 2009. . If you want to get a full essay! , parade it on our website: OrderEssay.net

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